Membrane Potential Responses Research Articles

A Potent Inhibitor of Phosphoinositide 3-Kinase (PI3K) and Mitogen Activated Protein (MAP) Kinase Signalling, Quercetin (3, 3', 4', 5, 7-Pentahydroxyflavone) Promotes Cell Death in Ultraviolet (UV)-B-Irradiated B16F10 Melanoma Cells

Ultraviolet (UV) radiation–induced skin damage contributes strongly to the formation of melanoma, a highly lethal form of skin cancer. Quercetin (Qu), the most widely consumed dietary bioflavonoid and well known inhibitor of phosphoinositide 3-kinase (PI3K) and mitogen activated protein (MAP) kinase signalling, has been reported to be chemopreventive in several forms of non-melanoma skin cancers. Here, we report that the treatment of ultraviolet (UV)-B-irradiated B16F10 melanoma cells with quercetin resulted in a dose dependent reduction in cell viability and increased apoptosis. The present study has brought out that the pro-apoptotic effects of quercetin in UVB-irradiated B16F10 cells are mediated through the elevation of intracellular reactive oxygen species (ROS) formation, calcium homeostasis imbalance, modulation of anti-oxidant defence response and depolarization of mitochondrial membrane potential (ΔΨM). Promotion of UVB-induced cell death by quercetin was further revealed by cleavage of chromosomal DNA, caspase activation, poly (ADP) ribose polymerase (PARP) cleavage, and an increase in sub-G1 cells. Quercetin markedly attenuated MEK-ERK signalling, influenced PI3K/Akt pathway, and potentially enhanced the UVB-induced NF-κB nuclear translocation. Furthermore, combined UVB and quercetin treatment decreased the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF). Overall, these results suggest the possibility of using quercetin in combination with UVB as a possible treatment option for melanoma in future.

Evaluación del potencial de membrana mitocondrial en espermatozoides humanos capacitados

Introducción: los espermatozoides humanos durante su viaje por el tracto reproductivo sufren el proceso de capacitación espermática, evento indispensable para la fecundación y que se ha relacionado con cambios en el potencial de membrana mitocondrial. Objetivos: evaluar el efecto de la capacitación espermática sobre el potencial de membrana mitocondrial en espermatozoides humanos. Materiales y métodos: se realizó un estudio in vitro en el cual se seleccionaron los espermatozoides puros de nueve voluntarios, los cuales se sometieron a condiciones capacitantes durante seis horas. Posteriormente, se evaluó el efecto de la capacitación sobre el estado del potencial de membrana mitocondrial usando el colorante DiOC6(3) por citometría de flujo. Resultados: de las nueve muestras seminales evaluadas cinco presentaron mayor intensidad media de fluorescencia en el potencial de membrana mitocondrial poscapacitación. Todas las muestras seminales mostraron parámetros normales y no presentaron cambios en la movilidad y la viabilidad espermática durante la selección y la capacitación espermática in vitro. Conclusiones: las modificaciones en el potencial de membrana mitocondrial en respuesta al proceso de capacitación demuestra que existen diferentes poblaciones espermáticas en el eyaculado humano; entender cuál está relacionada con el proceso reproductivo exitoso permitirá aumentar la probabilidad de embarazos.

Impairment of IKCa channels contributes to uteroplacental endothelial dysfunction in rat diabetic pregnancy.

Diabetes in rat pregnancy is associated with impaired vasodilation of the maternal uteroplacental vasculature. In the present study, we explored the role of endothelial cell (EC) Ca(2+)-activated K(+) channels of small conductance (SKCa channels) and intermediate conductance (IKCa channels) in diabetes-induced uterine vascular dysfunction. Diabetes was induced by injection of streptozotocin to second-day pregnant rats and confirmed by the development of maternal hyperglycemia. Control rats were injected with citrate buffer. Changes in smooth muscle cell intracellular Ca(2+) concentration, membrane potential, and vasodilation induced by SKCa/IKCa channel activators were studied in uteroplacental arteries of control and diabetic rats. The impact of diabetes on SKCa- and IKCa-mediated currents was explored in freshly dissociated ECs. NS309 evoked a potent vasodilation that was effectively inhibited by TRAM-34 but not by apamin. NS309-induced smooth muscle cell intracellular Ca(2+) concentration, membrane potential, and dilator responses were significantly diminished by diabetes; N-cyclohexyl-N-2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-4-pyrimidinamine (CyPPA)-evoked responses were not affected. Ca(2+)-activated ion currents in ECs were insensitive to paxilline, markedly inhibited by charybdotoxin (ChTX), and diminished by apamin. NS309-induced EC currents were generated mostly due to activation of ChTX-sensitive channels. Maternal diabetes resulted in a significant reduction in ChTX-sensitive currents with no effect on apamin-sensitive or CyPPA-induced currents. We concluded that IKCa channels play a prevalent role over SKCa channels in the generation of endothelial K(+) currents and vasodilation of uteroplacental arteries. Impaired function of IKCa channels importantly contributes to diabetes-induced uterine endothelial dysfunction. Therapeutic restoration of IKCa channel function may be a novel strategy for improvement of maternal uteroplacental blood flow in pregnancies complicated by diabetes.

Advantages and limitations of the use of optogenetic approach in studying fast-scale spike encoding.

Understanding single-neuron computations and encoding performed by spike-generation mechanisms of cortical neurons is one of the central challenges for cell electrophysiology and computational neuroscience. An established paradigm to study spike encoding in controlled conditions in vitro uses intracellular injection of a mixture of signals with fluctuating currents that mimic in vivo-like background activity. However this technique has two serious limitations: it uses current injection, while synaptic activation leads to changes of conductance, and current injection is technically most feasible in the soma, while the vast majority of synaptic inputs are located on the dendrites. Recent progress in optogenetics provides an opportunity to circumvent these limitations. Transgenic expression of light-activated ionic channels, such as Channelrhodopsin2 (ChR2), allows induction of controlled conductance changes even in thin distant dendrites. Here we show that photostimulation provides a useful extension of the tools to study neuronal encoding, but it has its own limitations. Optically induced fluctuating currents have a low cutoff (~70Hz), thus limiting the dynamic range of frequency response of cortical neurons. This leads to severe underestimation of the ability of neurons to phase-lock their firing to high frequency components of the input. This limitation could be worked around by using short (2 ms) light stimuli which produce membrane potential responses resembling EPSPs by their fast onset and prolonged decay kinetics. We show that combining application of short light stimuli to different parts of dendritic tree for mimicking distant EPSCs with somatic injection of fluctuating current that mimics fluctuations of membrane potential in vivo, allowed us to study fast encoding of artificial EPSPs photoinduced at different distances from the soma. We conclude that dendritic photostimulation of ChR2 with short light pulses provides a powerful tool to investigate population encoding of simulated synaptic potentials generated in dendrites at different distances from the soma.

Dissimilar responses of membrane potential (EM), permeability properties and respiration to cadmium and nickel in maize root cells

The short-term treatment with Cd2+ and Ni2+ triggered transient depolarization of transplasma membrane potential (EM) in the outer cortical root cells of two maize cultivars (cv. Premia and cv. Blitz), however, both metals changed the EM in a quantitatively different way. The magnitude and duration of EM depolarization were concentration dependent and were greater in the metal susceptible cv. Blitz. The highest EM depolarization was recorded with simultaneous application of Cd2+ + Ni2+ in both maize cultivars. The EM depolarization induced by Cd2+ or Cd2+ + Ni2+ but not Ni2+ alone was accompanied with a tremendous increase of membrane conductivity, but it was not accompanied with the effect of heavy metals (HM) on respiration. Simultaneous application of fusiccocin (FC) with Cd2+ or Cd2+ + Ni2+ during the EM depolarization, inability of FC to stop the depolarization by FC-enhanced proton extrusion and rapid restoration of EM, suggested a transient inhibition of the plasma membrane H+-ATPase by these toxic...

Using Nanoparticles to Control Cellular Membrane Potential

All cells generate an electrical potential (Vm) across their plasma membrane driven by a concentration gradient of charged ions. A typical resting membrane potential ranges from −40 to −70 mV, with a net negative charge on the cytosolic side of the membrane. Disruption of this electrochemical equilibrium causes Vm to become more positive or more negative relative to its resting state, referred to as “depolarization” or “hyperpolarization,” respectively. Changes in membrane potential have proven to be pivotal not only in normal cell cycle progression, but also in malignant transformation. Using polystyrene nanoparticles as a model system, we use a combination of fluorescence microscopy and flow cytometry to measure changes in membrane potential in response to nanoparticle binding to the plasma membrane. We find that cationic nanoparticles depolarize both CHO-K1 and HeLa cells. The cellular binding of anionic nanoparticles does not lead to a discernible trend in altered membrane potential. Maintenance of the resting membrane potential depends on the presence of two-pore-domain potassium “leak” channels, which allow for outward diffusion of potassium ions along their concentration gradient. Using an assay that tests the diffusion of ions through these potassium channels, we observe reduced permeability of the channels when cells are treated with nanoparticles. Based on a dynamical system model of the cell, we conclude that this loss of permeability likely results from physical blockage of the channel itself by the particle. Prevention of potassium ion efflux due to blocked channels causes accumulation of positive charge inside the cell, resulting in a depolarized membrane. By understanding the ways in which nanoparticles can be utilized to selectively generate cellular responses, we can begin to consider them as active species that may alter the very systems they are currently designed to probe.

Cellular binding of nanoparticles disrupts the membrane potential.

All cells generate an electrical potential across their plasma membrane driven by a concentration gradient of charged ions. A typical resting membrane potential ranges from -40 to -70 mV, with a net negative charge on the cytosolic side of the membrane. Maintenance of the resting membrane potential depends on the presence of two-pore-domain potassium "leak" channels, which allow for outward diffusion of potassium ions along their concentration gradient. Disruption of the ion gradient causes the membrane potential to become more positive or more negative relative to the resting state, referred to as "depolarization" or "hyperpolarization," respectively. Changes in membrane potential have proven to be pivotal, not only in normal cell cycle progression but also in malignant transformation and tissue regeneration. Using polystyrene nanoparticles as a model system, we use flow cytometry and fluorescence microscopy to measure changes in membrane potential in response to nanoparticle binding to the plasma membrane. We find that nanoparticles with amine-modified surfaces lead to significant depolarization of both CHO and HeLa cells. In comparison, carboxylate-modified nanoparticles do not cause depolarization. Mechanistic studies suggest that this nanoparticle-induced depolarization is the result of a physical blockage of the ion channels. These experiments show that nanoparticles can alter the biological system of interest in subtle, yet important, ways.

Zebrafish in auditory research: are fish better than mice?

Sensory inner and outer hair cells (IHCs and OHCs) in the mammalian organ of Corti mediate the perception of sound, and hair cells (HCs) in the vestibular system detect head movements. Their characteristic feature is the ‘hair bundle’ of actin-based stereocilia that are arranged in three rows, with increasing length toward the apical cell pole. Sound- or movement-induced deflection towards the longest row opens mechanoelectrical transduction (MET) channels, whereas movement in the opposite direction closes these ion channels. Accordingly, opening and closing of the MET channels transduce sound information and movement of the head into receptor potentials in auditory and vestibular hair cells, respectively. Depolarization induces calcium-dependent neurotransmitter release from specialized presynaptic structures in IHCs and vestibular HCs; the resultant EPSP modulates action potential firing in afferent nerves. In OHCs, receptor potentials produce somatic length changes that comprise the cochlear amplifier and underlie the high sensitivity and frequency selectivity of mammals. The response of the HC membrane potential to MET channel activation is tuned and characterized by a distinct profile of K+ channels in the basolateral membrane (Schwander et al. 2010).

Sound-induced hyperpolarization of hippocampal neurons

The hippocampus is involved in episodic memory, which is composed of subjective experiences in the multisensory world; however, little is known about the subthreshold membrane potential responses of individual hippocampal neurons to sensory stimuli. Using in-vivo whole-cell patch-clamp recordings from hippocampal CA1 neurons in awake mice, we found that almost all hippocampal neurons exhibited a hyperpolarization of 1-2 mV immediately after the onset of a sound. This large-scale hyperpolarization was unaffected by the duration or pitch of the tone. The response was abolished by general anesthesia and a surgical fimbria-fornix lesion.

Nonlinear properties of medial entorhinal cortex neurons reveal frequency selectivity during multi-sinusoidal stimulation.

The neurons in layer II of the medial entorhinal cortex are part of the grid cell network involved in the representation of space. Many of these neurons are likely to be stellate cells with specific oscillatory and firing properties important for their function. A fundamental understanding of the nonlinear basis of these oscillatory properties is critical for the development of theories of grid cell firing. In order to evaluate the behavior of stellate neurons, measurements of their quadratic responses were used to estimate a second order Volterra kernel. This paper uses an operator theory, termed quadratic sinusoidal analysis (QSA), which quantitatively determines that the quadratic response accounts for a major part of the nonlinearity observed at membrane potential levels characteristic of normal synaptic events. Practically, neurons were probed with multi-sinusoidal stimulations to determine a Hermitian operator that captures the quadratic function in the frequency domain. We have shown that the frequency content of the stimulation plays an important role in the characteristics of the nonlinear response, which can distort the linear response as well. Stimulations with enhanced low frequency amplitudes evoked a different nonlinear response than broadband profiles. The nonlinear analysis was also applied to spike frequencies and it was shown that the nonlinear response of subthreshold membrane potential at resonance frequencies near the threshold is similar to the nonlinear response of spike trains.

Less is more: minimal expression of myoendothelial gap junctions optimizes cell–cell communication in virtual arterioles

Dysfunctional electrical signalling within the arteriolar wall is a major cause of cardiovascular disease. The endothelial cell layer constitutes the primary electrical pathway, co-ordinating contraction of the overlying smooth muscle cell (SMC) layer. As myoendothelial gap junctions (MEGJs) provide direct contact between the cell layers, proper vasomotor responses are thought to depend on a high, uniform MEGJ density. However, MEGJs are observed to be expressed heterogeneously within and among vascular beds. This discrepancy is addressed in the present study. As no direct measures of MEGJ conductance exist, we employed a computational modelling approach to vary the number, conductance and distribution of MEGJs. Our simulations demonstrate that a minimal number of randomly distributed MEGJs augment arteriolar cell-cell communication by increasing conduction efficiency and ensuring appropriate membrane potential responses in SMCs. We show that electrical coupling between SMCs must be tailored to the particular MEGJ distribution. Finally, observation of non-decaying mechanical conduction in arterioles without regeneration has been a long-standing controversy in the microvascular field. As heterogeneous MEGJ distributions provide for different conduction profiles along the cell layers, we demonstrate that a non-decaying conduction profile is possible in the SMC layer of a vessel with passive electrical properties. These intriguing findings redefine the concept of efficient electrical communication in the microcirculation, illustrating how heterogeneous properties, ubiquitous in biological systems, may have a profound impact on system behaviour and how acute local and global flow control is explained from the biophysical foundations.

Regulation of KCa2.3 and endothelium-dependent hyperpolarization (EDH) in the rat middle cerebral artery: the role of lipoxygenase metabolites and isoprostanes.

Background and Purpose. In rat middle cerebral arteries, endothelium-dependent hyperpolarization (EDH) is mediated by activation of calcium-activated potassium (KCa) channels specifically KCa2.3 and KCa3.1. Lipoxygenase (LOX) products function as endothelium-derived hyperpolarizing factors (EDHFs) in rabbit arteries by stimulating KCa2.3. We investigated if LOX products contribute to EDH in rat cerebral arteries.Methods. Arachidonic acid (AA) metabolites produced in middle cerebral arteries were measured using HPLC and LC/MS. Vascular tension and membrane potential responses to SLIGRL were simultaneously recorded using wire myography and intracellular microelectrodes.Results. SLIGRL, an agonist at PAR2 receptors, caused EDH that was inhibited by a combination of KCa2.3 and KCa3.1 blockade. Non-selective LOX-inhibition reduced EDH, whereas inhibition of 12-LOX had no effect. Soluble epoxide hydrolase (sEH) inhibition enhanced the KCa2.3 component of EDH. Following NO synthase (NOS) inhibition, the KCa2.3 component of EDH was absent. Using HPLC, middle cerebral arteries metabolized 14C-AA to 15- and 12-LOX products under control conditions. With NOS inhibition, there was little change in LOX metabolites, but increased F-type isoprostanes. 8-iso-PGF2α inhibited the KCa2.3 component of EDH.Conclusions. LOX metabolites mediate EDH in rat middle cerebral arteries. Inhibition of sEH increases the KCa2.3 component of EDH. Following NOS inhibition, loss of KCa2.3 function is independent of changes in LOX production or sEH inhibition but due to increased isoprostane production and subsequent stimulation of TP receptors. These findings have important implications in diseases associated with loss of NO signaling such as stroke; where inhibition of sEH and/or isoprostane formation may of benefit.

Sonoporation-Induced Depolarization of Plasma Membrane Potential: Analysis of Heterogeneous Impact

Disrupting plasma membrane integrity would inevitably promote anomalous ion fluxes across the membrane and thereby upset the trans-membranous potential. In this article, we report new findings on how sonoporation as a physical membrane perforation strategy would lead to different forms of plasma membrane potential disruption. Our investigation was conducted with a customized fluorescence imaging platform that enabled live monitoring of plasma membrane potential in relation to individual sonoporation events triggered on HeLa cervical cancer cells. Sonovue microbubbles were used as sonoporation agents (added at a 4:3 cell-to-bubble ratio), and they were activated by 1-MHz pulsed ultrasound with 0.35-MPa peak negative pressure, 20-cycle pulse duration, 20-Hz pulse repetition frequency and 1-s total exposure duration. Results indicate that the plasma membrane potential response was heterogeneous among sonoporated cells: (i) membrane potential of irreversibly sonoporated cells was permanently depolarized; (ii) reversibly sonoporated cells exhibited either transient or sustained membrane depolarization; (iii) intact cells adjacent to sonoporated ones underwent transitory membrane depolarization. These findings effectively serve to substantiate the causal relationship between sonoporation and plasma membrane potential.

Strabismus Disrupts Binocular Synaptic Integration in Primary Visual Cortex

Visual disruption early in development dramatically changes how primary visual cortex neurons integrate binocular inputs. The disruption is paradigmatic for investigating the synaptic basis of long-term changes in cortical function, because the primary visual cortex is the site of binocular convergence. The underlying alterations in circuitry by visual disruption remain poorly understood. Here we compare membrane potential responses, observed via whole-cell recordings in vivo, of primary visual cortex neurons in normal adult cats with those of cats in which strabismus was induced before the developmental critical period. In strabismic cats, we observed a dramatic shift in the ocular dominance distribution of simple cells, the first stage of visual cortical processing, toward responding to one eye instead of both, but not in complex cells, which receive inputs from simple cells. Both simple and complex cells no longer conveyed the binocular information needed for depth perception based on binocular cues. There was concomitant binocular suppression such that responses were weaker with binocular than with monocular stimulation. Our estimates of the excitatory and inhibitory input to single neurons indicate binocular suppression that was not evident in synaptic excitation, but arose de novo because of synaptic inhibition. Further constraints on circuit models of plasticity result from indications that the ratio of excitation to inhibition evoked by monocular stimulation decreased mainly for nonpreferred eye stimulation. Although we documented changes in synaptic input throughout primary visual cortex, a circuit model with plasticity at only thalamocortical synapses is sufficient to account for our observations.

Abstract 538: Prolonged Functional Sympathetic Efferent Denervation Following Ablation of the Renal Nerves in SHR

Renal denervation (RDN) has been shown to be an effective and safe treatment for human resistant hypertension and is considered one of the most promising advances in the field of hypertension. However, the mechanisms mediating the sustained decrease in arterial pressure in response to RDN in humans are unknown. This issue is particularly perplexing, given reports that the depressor effect of RDN in animal models is short lived. The current dogma in the literature is that overtime the efferent sympathetic nerves regrow and so the longevity of the depressor effect of RDN in humans cannot be ascribed to the absence of efferent renal sympathetic nerve activity. We hypothesize that the renal sympathetic nerves regenerate following RDN, and arrive at the target tissues, but the functional response is not fully restored and this contributes to the long-term depressor effect of RDN. Our aim was to examine mean arterial pressure (MAP) and vascular function in spontaneously hypertensive rats (SHR) following RDN. In 10 week old male SHR radiotelemetry probes were implanted to measure MAP. At 12 weeks of age, following 3 days basal MAP recording, sham (n=5) or bilateral RDN (n=5) surgery was performed. MAP was recorded until 24 weeks of age (12 weeks post-RDN). Renal lobar arteries were mounted on a myograph and membrane potential and tension responses to perivascular nerve stimulation assessed. Perivascular nerve stimulation with single pulses at increasing voltage evoked excitatory junction potentials (Ejps) of increasing amplitude in SHR-sham. Ejp amplitude was markedly reduced in arteries from SHR-RDN (P<0.01). Repetitive stimulation (1-8Hz, 5s) evoked slow depolarisation and contraction. At 5Hz 5s, slow depolarisation was 12±2 mV in SHR-shams and 7±2 mV in SHR-RDN (P<0.01), and contractions were smaller than SHR-sham (10 ± 3% vs 22 ± 2% HiK; P<0.01). In conclusion, smaller Ejps suggest that close contacting neuroeffector junctions are not reformed and blunted vasoconstrictor responses indicate neuro-vascular function has only partially returned 12 weeks post-RDN (Fig. 2). Our finding of continued ‘functional’ denervation is compatible with our data that RDN causes a sustained decrease in arterial pressure in SHR.

Predicting the firing phase of an oscillatory neuron from its impedance profile

The activity phase of a neuron in an oscillatory network often determines what the neuron codes [1,2]. We are interested in understanding the effect of subthreshold factors that influence this activity phase. Here we develop a first-order approximation of the activity phase of a neuron receiving oscillatory input using its subthreshold impedance profile. A neuron's subthreshold membrane potential response to sinusoidal current input with frequency f is sinusoidal (to first order) with amplitude and phase-shift approximated by the impedance value at f:Zf=Zfeiφf. If a neuron receives suprathreshold oscillatory input at frequency f, the resulting change in membrane potential can be approximated with a similar amplitude and phase-shift up to the time point tspike where spike threshold is reached. This results in the following simple equation: Vm(tspike)=Vrest+AinZfsin(2πf⋅tspike+φf)=Vthresh where Ain is the amplitude of the input current. Assuming the reference time point of t0 = 0 in the cycle, the spike phase can be approximated as φspike=f⋅tspike=12πarcsinVthresh-VrestAinZf-φf. This approximation is valid so long as the argument of arcsin is <1 in absolute value. As proof of principle, we used the impedance profile of a model neuron exhibiting subthreshold resonance to approximate the spike phase with a given preset spike threshold (Figure A). We also used this approximation to predict the phase of the first spike in bursting PY neurons in the crab pyloric CPG, when synaptically isolated and subjected to a sinusoidal current input at different frequencies (0.1-4 Hz; see [3]for method). The PY impedance was measured from its subthreshold response. To our knowledge this method, despite its simplicity, has not been previously used for the approximation of spike phase using the impedance profile. The usefulness of this approximation is that changes in membrane properties due to network activity or neuromodulation can be readily measured in the impedance profile and this knowledge can be used to predict how the neuron changes its response phase during network activity. There are three sources of error in this approximation. First, the membrane impedance in biological neurons is nonlinear and does not scale linearly with the amplitude of the input current, especially when the neuron transitions from sub- to suprathreshold activity. However, the shift in membrane impedance can be tracked in both model and biological neurons. Second, spike threshold is dependent on input frequency. This dependence can also be tracked. Third, because of nonlinearities, the membrane potential response of a neuron is not perfectly sinusoidal. As seen in Figure ​Figure1B,1B, despite these (nonlinear) sources of error, our method can provide a good approximation of spike phase. We are in the process of estimating spike phase in response to synaptic inputs arriving at a fixed frequency. Figure 1 A. The spike phase of a resonate-and-fire model predicted from the impedance profile. B. The spike phase of the PY neuron subject to a ZAP input (inset) predicted from its subthreshold response (green).

Shot Noise analysis of subthreshold membrane potential activity in neurons

Shot noise is one of the most extensively harnessed phenomena used to model different aspects of neural activity in neuroscience. Shot noise processes [1] display very characteristic fluctuations that correspond to the sum of causal system responses to unitary event arrivals that follow a Poisson distribution. In general, applications of shot noise [2-4] have been confined to the so-called Campbell Limit corresponding to steady-state regime, or to specific post-synaptic responses that make the process Markovian or with the assumption of homogeneous Poisson event arrivals. In this work we go beyond this context to show that the shot-noise formalism can be used to analyze the synaptic activity of neurons beyond the Campbell limit, for any type of bounded post-synaptic response function and non-homogeneous Poisson event arrivals. This enhancement of technique allows for two main applications. 1) Computation of the full density for current synapses and statistical moments for conductance synapses. This yields a statistical description of the subthreshold membrane potential. This includes considering distributions on the afferent populations over biological parameters such as peak intensity, number of independent channels, or dynamical parameters such as distribution of incoming firing rate or variable incoming firing rate. The picture obtained is of particular interest for various applications such as calculating the transfer function. 2) Under stationarity assumptions, this technique offers the potential to reverse the moments of membrane potential integration. This offers a candidate method to infer characteristics of the synaptic activity (and therefore of the network activity) from the analysis of the subthreshold variations of the membrane potential. This application is of particular interest for the analysis of intracellular recordings in vivo. We illustrate these techniques by comparing analytic expressions with numerical estimates, as well as to neurons subject to a large range of synaptic activity (including low release rate far from the diffusion limit). In conclusion, due to their flexibility as a modeling tool, shot noise analysis appears as a very useful tool in computational neuroscience to model post-synaptic and sub-threshold membrane potential responses to incoming network activity, and also as a tool for electrophysiological analysis of network activity from intracellular recordings.

Sublinear binocular integration preserves orientation selectivity in mouse visual cortex

Inputs from the two eyes are first combined in simple cells in the primary visual cortex. Consequently, visual cortical neurons need to have the flexibility to encode visual features under both monocular and binocular situations. Here we show that binocular orientation selectivity of mouse simple cells is nearly identical to monocular orientation selectivity in both anesthetized and awake conditions. In vivo whole-cell recordings reveal that the binocular integration of membrane potential responses is sublinear. The sublinear integration keeps binocularly-evoked depolarizations below threshold at non-preferred orientations, thus preserving orientation selectivity. Computational simulations based on measured synaptic conductances indicate that inhibition promotes sublinear binocular integration, which are further confirmed by experiments using genetic and pharmacological manipulations. Our findings therefore reveal a cellular mechanism for how visual system can switch effortlessly between monocular and binocular conditions. The same mechanism may apply to other sensory systems that also integrate multiple channels of inputs.

Leptin regulates surface expression of β ‐cell ATP‐sensitive potassium channels via an AMPK/PKA/Cofilin signaling cascade

An adipoinsular axis is a dual hormonal feedback loop in which insulin produced by pancreatic β‐cells increases leptin production from adipose tissue and leptin released from adipose tissue in turn down regulates insulin secretion. Leptin has been shown to reduce insulin secretion at least in part by increasing KATP channel conductance, a key regulator of β‐cell membrane potential in response to glucose metabolism. However, the mechanism by which leptin increases KATP currents is unknown. We report here that leptin increases KATP channel expression in the β‐cell plasma membrane. Using a combination of pharmacological and molecular genetics approach, we show that this effect is mediated by a signaling cascade involving activation of AMPK via phosphorylation at Thr‐172 and activation of PKA, which then leads to phoshorylation of the actin binding protein cofilin and actin depolymerization. Imaging and surface biotinylation pulsechase experiments show that increased surface KATP channel abundance in response to leptin is primarily a consequence of increased KATP channel trafficking to the cell surface and that this trafficking event is tightly correlated with actin depolymerization. Taken together, our findings suggested that leptin inhibit insulin secretion by recruiting KATP channels to the β‐cell plasma membrane via a AMPK/PKA/cofiln/F‐actin depolymerization signaling mechanism. (This work was supported by NIDDK057699.)

Odorant-induced membrane potential depolarization of AIY interneuron in Caenorhabditis elegans

Although some interneurons in C. elegans have been shown to have unusual region-specific Ca2+ dynamics, the region-specific Ca2+ and membrane potential response properties of these neurons are largely unknown due to technical limitations. In this report, we focused on one of these neurons, AIY interneuron, where Ca2+ dynamics have been detected only in neurites, and not the soma, during odor and temperature stimulation to determine whether membrane potential and Ca2+ are region-specific dynamics and distinct from one another. To visualize voltage change both in the soma and neurites of AIY, we used voltage-sensitive fluorescent protein (VSFP) 2.42. First, we confirmed that the sensor protein worked correctly in C. elegans by depolarizing AIY interneuron with high concentrations of KCl. Next, we observed membrane potential depolarization during odor (isoamyl alcohol) stimulation in both neurites and the soma. Additionally, depolarization of membrane potential with direct application of high KCl induced a Ca2+ increase in the soma. From these results, we conclude that membrane potential behavior and Ca2+ dynamics in AIY differ in its subcellular regions and that VSFP2.42 can be a useful tool for studying information processing in single neurons.